Thumbnail Image

Thesis

Browse

2014 - 20173
Reset filters
Subject: Veterinary Physiology × End date: 2018 × Start date: 2014 ×
Reset filters

Search Results

Now showing 1 - 3 of 3
  • Thumbnail Image
    ThesisItemOpen Access
    LEPTIN RECEPTOR EXPRESSION IN CULTURED OVARIAN FOLLICLES OF SHEEP
    (SRI VENKATESWARA VETERINARY UNIVERSITY TIRUPATI - 517 502. (A.P.) INDIA, 2017-05) SRAVANI PRAGNA, K; Siva Kumar, A.V.N(MAJOR); Rambabu Naik, B; Punya Kumari, B
    ABSTRACT : The expression of Leptin receptor (LepR) mRNA and protein was studied in sheep using Real time quantitative PCR and Immunohistochemistry in the cumulus cells and oocytes from : (i) In vivo grown preantral, early antral, antral, large antral follicles and Cumulus Oocyte Complexes (COCs) obtained from large antral follicles and subjected to 24h of In Vitro Maturation ( IVM) and (ii) Preantral follicles (PFs’) exposed to TCM199B, TCM199BL (10ng/mL) and TCM199BHGL (10ng/mL) media for 3min, cultured in vitro for two, four or six days and subsequently matured in vitro for 24h in respective cultures separately. LepR was observed at all stages of in vivo grown and in vitro cultured ovarian follicles in both cumulus cells and oocytes. Expression levels and patterns of LepR mRNA in PFs’ cultured in TCM199BHGL was similar to in vivo in all the stages except in cumulus cells from COCs after in vitro maturation for 24h, where the expression was significantly higher which was a positive effect. The expression of LepR in cumulus cells among the three different in vitro culturing conditions was significantly higher in PFs’ cultured in TCM199BHGL medium. In the oocytes, the Leptin receptor expression was similar or significantly higher in PFs’ cultured in TCM199BHGL medium than in vivo grown follicles except in PFs’ exposed to 3min stage which could suggest the synergistic action of growth factors and hormones with Leptin. Immunohistochemistry studies revealed highest intensity of Leptin receptor protein expression in the oocytes. Mild to moderate staining was observed in cumulus cells with good intensity in large antral stages. This protein expression coincided with gene expression. From our studies it is concluded that Leptin along with other growth factors and hormones supplementation stimulated the expression of Leptin receptor mRNA and protein on par with that of in vivo /even higher than that of in vivo condition.
  • Thumbnail Image
    ThesisItemOpen Access
    INFLUENCE OF LEPTIN ON THE EXPRESSION OF p53 and BAK GENES IN CULTURED OVARIAN FOLLICLES IN SHEEP
    (SRI VENKATESWARA VETERINARY UNIVERSITY TIRUPATI - 517 502. (A.P.) INDIA, 2015-11) PRABHAVATHI, T; RAO, V.H(MAJOR); SIVA KUMAR, A.V.N; VEERABRAHMAIAH, K
    ABSTRACT : The present study was conducted to know the influence of Leptin on the expression of p53 and BAK (apoptotic) genes in the sheep preantral follicles (PFs’) cultured in vitro. Ovaries of sheep were collected from the local slaughter house. Intact PFs’ (250-400μm) were isolated and placed individually in 20μl droplets of standard medium (TCM 199 B + 2 μg/ml FSH, 0.1μg/ml T4 + 10 ng/ml IGF-I + 1 mIU/ml of GH) supplemented with Leptin (10ng/ml) for 2, 4 or 6 days. Quantitative expression of Test genes (p53 and BAK) and Reference genes (18s RNA, RPLPO, HPRT1) were studied in the cumulus cells and oocytes isolated at different developmental stages (preantral, early antral, antral and large antral follicles) of the in vivo and in vitro grown follicles. The entire experiment was repeated twice. Duplicate samples of cDNA from each replicate of the experiment were subjected to relative-RT-qPCR. The pattern of expression of p53 gene in the cumulus cells from ovarian follicles grown in vivo and cultured in Leptin was similar from preantral to large antral follicle stage. In the oocytes, however, such similarity was restricted up to the antral follicle stage only. It is concluded that Leptin in culture medium mimiced the pattern of p53 expression in in vitro as in in vivo to some extent. The BAK gene expression was undetected in the cumulus cells and oocytes isolated from all the stages of both in vivo and in vitro developed ovarian follicles. Accordingly it is concluded that (i) Leptin has no influence on the expression of BAK gene in cultured ovarian follicles in sheep (ii) BAK may not be an important regulator of apoptosis in ovarian follicles and (iii) p53 plays a relatively more significant role than BAK in the regulation of apoptosis in ovarian follicles.
  • Thumbnail Image
    ThesisItemOpen Access
    INFLUENCE OF LEPTIN ON IN VITRO DEVELOPMENT OF PREANTRAL FOLLICLES OF SHEEP
    (SRI VENKATESWARA VETERINARY UNIVERSITY TIRUPATI - 517 502. (A.P.) INDIA, 2014-11) Siva Kumar, A.V.N.; Rao, V.H(MAJOR); Punya Kumari, B
    ABSTRACT : The role of human and ovine leptin on in vitro development of preantral follicles (PFs’) isolated from sheep ovaries was studied. Experiment I, was conducted to standardize the optimum concentration of human leptin for the growth of PFs’. The parameters set for the evaluation of the growth of PFs’ were increase in diameter, proportion of PFs’ exhibiting growth, antrum formation and maturation of oocytes to M II stage. All these parameters were higher in the leptin (10ng/ml) supplemented medium. Experiment II was conducted with the optimum level of leptin (10ng/ml) derived in experiment I to evaluate the effect of human or ovine leptin @10ng/ml in combination with various growth factors viz., TCM 199 B with FSH, T4, IGF-I and GH. Two controls viz., TCM 199 B and standard medium i.e., TCM 199 B supplemented with FSH (2ug/ml), T4 (1μg/ml), IGF-I (10ng/ml) and GH (1mIU/ml) were used. In the standard medium supplemented with human leptin, the proportion of PFs’ exhibiting growth (66.14±3.9), average increase in the diameter (11.1±0.87 μ), antrum formation (51.5±5.33) and maturation of oocytes to M II stage (24.1±3.7) were higher compared to the standard medium and standard medium supplemented with ovine leptin (10ng/ml). But there was no significant difference between the standard medium and the standard medium supplemented with ovine/human leptin. It is concluded that (i) the optimum dose of leptin for the growth of sheep PFs’ in vitro was 10ng/ml and (ii) human or ovine leptin supported similar growth of PFs’ cultured in the standard medium.